The distribution of the activity of brain hexokinase between soluble and mitochondrial forms is at least partially governed by metabolic activity, increased amounts of the particulate enzyme being observed during times of increased glycolytic activity. These changes may be involved in regulation of activity and are thought to result from alteration in intracellular levels of various metabolites (e.g., glucose-6-P, Pi) previously shown to influence this distribution in vitro. The importance of the primary structure and conformation of the enzyme, and the role of divalent cations and various constituents of the outer mitochondrial membrane (e.g., lipids, proteins, etc.) in this specific membrane-enzyme interaction will be investigated. Localization of hexokinase at the electron microscopic level will be attempted using immunoperoxidase techniques. This methodology will be of interest: a) in determining the distribution of hexokinase binding sites on mitochondria (random vs. specific localizations), b) in assessing the soluble-mitochondrial distribution of the enzyme in different cell-types, or as it is influenced by different metabolic states. BIBLIOGRAPHIC REFERENCES: Manthorpe, C.M., Jr., Nettleton, D.O., and Wilson, J.E. (1976) Purification of Rat Brain Mitochondria: A New Procedure. J. Neurochem. 27, 1547-1549. Wilson, J.E. (1976) Applications of Blue Dextran and Cibacron Blue F3GA in Purification and Structural Studies of Nucleotide-Binding Enzymes. Biochem. Biophys. Res. Commun. 72, 816-823.